Journal of clinical virology plus最新文献

The SARS CoV-2 D614G variant circulated in Cuba in 2020. New viral variants were detected after the opening of the border in November 2020. We show the results of the genomic surveillance in Cuba from December 28, 2020, to September 28, 2021 and their relationship to the epidemiological situation in the country. A total of 1,406 nasopharyngeal exudates from COVID-19 patients were processed for RNA extraction and the 1836 bp fragment of the spike gene was amplified and sequenced. The mutations present were determined using the GISAID database. Prevalence ratios were estimated by fitting Poisson univariate and multivariate regression models to investigate associations between SARS-CoV-2 variant group (VOC, non-VOC) and disease outcome. Seventeen genetic variants were detected including VOC Alpha, Beta, Gamma and Delta, one variant of interest (VOI) (Lambda) and two previous VOI (A.2.5.1 and Zeta/P.2). Beta (34.77%), Delta (24.89%) and D614G (19%) variants were the most frequently detected. By June, Delta increased in frequency, displacing Beta. Disease severity increased significantly with age and VOC (PR =1.98, IC 95%: 1.33–3.05, p <0.05). Genomic surveillance allowed us to identify the upsurge of novel variants. Coinciding with the higher epidemic period, multiple variants were co-circulating. Although we cannot rule out that failure in the transmission containment measures occurred, the increase in the number of cases associated with the circulation of several variants, particularly the Beta and Delta variants is highly suggestive. A greater association of Beta variant with clinical severity and Delta variant with a greater transmissibility was observed.

Background

Over 296 million people worldwide are living with chronic Hepatitis B (CHB) infection who require monitoring of viral activity and disease progression. Serum HBV RNA is a promising new biomarker in CHB management. No standardized method for serum HBV RNA quantification has been established.

Objectives

To develop and validate, both analytically and clinically, a 3′ RACE RT-qPCR assay for quantification of serum HBV RNA.

Study design

The 3′ RACE RT-qPCR method was developed using published primers. The analytical limit of detection and quantification, linearity, inter- and intra-assay repeatability, and clinical specificity and sensitivity were evaluated using synthetic pre-genomic RNA and specimens from various patient populations. Intra- and inter-laboratory ring trials involving three laboratories were completed.

Results

The 3′ RACE RT-qPCR assay dynamic range is 25 to 10⁸ copies/µL of synthetic RNA. Theoretical and measured serum HBV RNA quantities from a serially diluted sample showed excellent linearity (R²=0.9795). The inter- and intra-assay repeatability were 94.91% and 95.12%, respectively. Clinical specificity and sensitivity were 100% and 90%, respectively, in treated patients. Inter-laboratory analysis demonstrated moderate to high agreement among participating laboratories (κ = 0.581 to 0.867). High agreement was also observed between both operators participating in the intra-laboratory evaluation (κ = 0.867).

Conclusions

Our methodology for the quantification of serum HBV RNA is specific and repeatable and employs a biologically relevant RNA standard suitable for medium throughput laboratories. Method standardization is required to facilitate the comparison of studies and better understand the clinical role of this novel biomarker.

In 2010 and 2012, between June and October (the peak period of enterovirus circulation in Nigeria), 59 archived enterovirus isolates (recovered as part of the Acute Flaccid Paralysis surveillance program) were randomly selected and typed in this study. All isolates were subjected to RNA extraction, cDNA synthesis and VP1 amplification (nested) and sequencing. Of the 59 isolates, 45 (21 and 24 of the isolates from 2010 to 2012, respectively) had the expected band size (∼350 bp). Forty-three (43) (19 and 24 from 2010 to 2012, respectively) of the amplicons generated were successfully identified and all typed as Enterovirus species B (EV-B) members belonging to twenty (20) different types. The most commonly detected were Echovirus 11 (E11) (9 isolates) and E30 (4 isolates). Many EV-B clades detected show evidence of cross-border transmission in humans within Africa and international spread. Evidence of possible zooanthroponosis/anthropozoonosis between human and nonhuman primates in one EV-B lineage is also reported.

The Omicron emerged in November 2021 and became the predominant SARS-CoV-2 variant globally. It spreads more rapidly than ancestral lineages and its rapid detection is critical for the prevention of disease outbreaks. Antigen tests such as immunochromatographic assay (ICA) and chemiluminescent enzyme immunoassay (CLEIA) yield results more quickly than standard polymerase chain reaction (PCR). However, their utility for the detection of the Omicron variant remains unclear. We herein evaluated the performance of ICA and CLEIA in saliva from 51 patients with Omicron and 60 PCR negative individuals. The sensitivity and specificity of CLEIA were 98.0% (95%CI: 89.6–100.0%) and 100.0% (95%CI: 94.0–100.0%), respectively, with fine correlation with cycle threshold (Ct) values. The sensitivity and specificity of ICA were 58.8% (95%CI: 44.2-72.4%) and 100.0% (95%CI: 94.0–100.0%), respectively. The sensitivity of ICA was 100.0% (95%CI: 80.5–100.0%) when PCR Ct was less than 25. The Omicron can be efficiently detected in saliva by CLEIA. ICA also detects high viral load Omicron using saliva.

In the current study, an one pot quadruplex RT-qPCR assay was established and comprehensively evaluated. The assay's limit of detection could reach as low as 10¹ copies/reaction, with good repeatability profile and no cross-reaction with other respiratory pathogens. During clinical evaluation both by blinded samples and real clinical samples, the assay exhibited a 100% coincidence rate with individual commercial RT-qPCR assays. To the best of our knowledge, this is the first report on the simultaneous subtyping influenza A virus into H1, H3, N1, and N2 by one pot quadruplex RT-qPCR assay, which could improve the preparedness for future influenza outbreaks.

Background

Human papillomavirus (HPV) is known to be associated with both cervical and oropharyngeal cancers, yet little are known about the co-occurrence of similar HPV subtypes in these subsites. With the changing sexual behaviour it is necessary to evaluate the risk of a woman with abnormal cervix developing cancer of the oropharynx. A study evaluating HPV infection in both oral mucosa and the cervix is therefore needed to understand the co-occurrence of HPV infection in 2 different sites

Methods

Oral mucosa and cervical wash specimens were collected from a total of 100 women aged 20-60 years who visited for routine Pap test. DNA was extracted and then subjected to an in house real-time PCR and conventional nested PCR for the detection high risk HPV 16/18 subtypes. A limit of detection was established using serially diluted HPV positive DNA from cell lines and the sensitivity and specificity of both the assays were evaluated

Results

The prevalence of HPV-16/18 in the cervical samples (19.7% vs 6.2%) and the oral mucosa (16.6% vs nil) were higher as detected by our in house real-time PCR in comparison to the conventional nested PCR. Oral HPV infection was found in 25% of the women with SCC of the cervix

Conclusion

Due to the changing sexual behaviour there is a possibility that women with HPV-related cervical lesions can easily transfer the virus to the partner or even self-inoculate which might lead to the development of lesions in the head & neck turning into cancer.

Background

There is a strong association between high-level BK viraemia and the development of BK virus-associated nephropathy. Therefore, understanding the kinetics of BK virus replication following transplantation would be helpful in predicting high-level viraemia and the development of BKVAN.

Objectives

This study was aimed to assess the incidence of BKV infection and the BK viral kinetics among renal transplant recipients in their first year following the transplant.

Methods

This study was a retrospective descriptive study involving renal transplant recipients treated at National Hospital, Kandy, Sri Lanka. The patients were recruited from January 2018 to December 2019 and their demographic details and quantitative BKV PCR assay results were analysed during the first year following transplantation.

Results

297 patients were transplanted during the study period wherein, males predominated with 78.1% and 43.67 years old being the mean age (IQR: 35.50–53.0). The cumulative incidence of BK viraemia was 24.9% (n = 74). The mean age of the cohort with BK viraemia was 44.4 years (IQR: 36–54). Twenty percent (n = 13) of the female population had BKV viraemia, while 26.29% (n = 61) of the male population. Among patients with BK viraemia 12 had significant viraemia of 104IU/ml giving a cumulative incidence of significant viraemia was 4.04% and 10 of them (83.33%) had low levels of viraemia before showing significant viraemia.

Conclusion

The presence of low-level viraemia in 83% of patients with significant viraemia proves the value of early routine screening to predict the development of significant BK viraemia and to take preventive measures.

Background

Chronic kidney disease (CKD) remains an important comorbid condition in people living with HIV. However, data in children living with HIV/AIDS (CLWHA) in sub-Saharan Africa is limited. We sought to establish the prevalence and identify risk factors of CKD among CLWHA in SSA.

Methods

This was a retrospective chart review across five SSA countries HIV/AIDS care sites, March 2000 and June 2016.

Results

4,859 children with at least two clinic visits were enrolled in the study. The median age at the first clinic visit was 5.7 (IQR; 2.5, 9.5) years, and median follow-up time was 22.6 (IQR 9.8, 46.1) months. 11.2% CLWHA had an eGFR of <60 mL/min/1.73m² on at least one clinic visit. The prevalence of CKD was 1.6%. In a multivariable Poisson regression analysis, CKD was associated with severe immunosuppression, incident rate ratio (IRR) 2.69 (95% CI, 1.11, 6.51). Risk of CKD decreased with increasing age (IRR 0.51 (95% CI, 0.39, 0.67). There was no association between CKD and ART regimen.

Conclusion

CKD was not as prevalent as previously reported in children in other studies. Kidney function monitoring should be incorporated into the pediatric HIV care monitoring guidelines to allow for better evaluation of kidney disease in CLWHA.